Recognition of foreign antigens by mammalian cells can be mediated by a set of innate immune receptors called Toll-like receptors (TLRs). TLRs recognize conserved patterns derived from microbial pathogens identified as pathogen-associated molecular patterns (PAMPs) (Barton et al., Science 300:1524-1525, 2003). Interaction of a TLR with a PAMP results in a signaling cascade involving NF-κB activation and transcription of cytokine gene expression. Ten human toll-like receptors and five TLR adaptor proteins have been identified.
TLRs are able to expand their repertoire of ligands by forming homo- or heterodimers as well as binding different adaptor proteins. For example, TLR3 binds dsRNA, an intermediate in viral replication. TLR3 also interacts with PolyI:C, a synthetic dsRNA analog, and mRNA from necrotic cells. Activation of TLR3 leads to the secretion of Type I interferons, which are important in the control of viral infection. A full-length human TLR3 amino acid sequence and encoding polynucleotide sequence are shown in SEQ ID NOs: 1 and 2, respectively. TLRs TLR7, TLR8, and TLR9 also have nucleic acid ligands; activation of these TLRs can also lead to interferon secretion.
Type I interferons trigger signaling cascades to activate a set of immediate early-response genes (IFN-stimulated genes or ISGs) and have proven useful in the clinic. The resulting antiviral activities include mRNA translation inhibition, RNA editing, and RNA degradation (Samuel et al., Clin Microbiol Rev 14:778-809, 2001). Currently, a combination therapy of pegylated interferon and the broad-spectrum antiviral compound ribavirin is being used to treat hepatitis C infection (Manns et al., Lancet 358:958-965, 2001).
The critical anti-viral role of Type I IFNs is further demonstrated by the evolution of viral resistance mechanisms to inhibit the production of Type I IFNs by infected host cells. For example, the NS1 protein of influenza antagonizes IRF-3 activation and IFNβ production (Donelan et al., J Virol 78: 11574-11582, 2004) and the A52R poxvirus protein associates with IRAK2 and TRAF6 to block signaling downstream of TLR3 (Harte et al., J Exp Med, 197:343-351, 2003). Thus, therapies based on triggering TLR activation or enhancing TLR-mediated signaling pathways increase endogenous IFNα/β production and assist the host in the control of acute viral infections.
The use of TLR agonists to modulate the outcome of an immune response is currently being investigated for therapeutic use (O'Neill, Curr Opin Pharm 3:396-403, 2003; Schetter et al., Curr Opin Drug Discov Devel 7:204-210, 2004). For example, CpG oligodinucleotides (ODN), a TLR9 ligand, are capable of stimulating the production of type I IFN and a TE1 response (Krieg, Annu Rev Immunol 20:709-60, 2002), a finding that suggests the possible use of CpG ODN not only as a vaccine adjuvant but also for the treatment and or prevention of diseases that necessitate a potent TH1 response. Another example is the synthetic TLR7 agonist imiquimod, an approved agent for the treatment of genital warts; its protective effect is thought to be mediated through the stimulation of inflammatory cytokines such as IFNα, TNFα and IL-1β (Saunder, J Amer Acad Derm 43: S6-S11, 2000). Overall, these findings show that TLR agonists are a novel class of immunomodulatory agents with the potential of having a significant therapeutic benefit.
Thus, a need exists for the identification of novel immunomodulatory agents that potentiate the effect of TLR agonists. Such novel TLR-based therapies are expected to have an advantage of providing a sustained immune response with less frequent dosing regimens.